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O-Glycoprotease is a highly specific protease that cleaves the peptide bonds of a glycoprotein or glycopeptide immediately N-terminal to a serine or threonine residue containing a mucin-typeO-linked glycan with or without sialylation.
Enables O-glycosylation site mapping and O-glycan structural determination
Efficiently cleaves glycoproteins with or without sialic acid, no neuraminidase treatment necessary
Recombinant enzyme with no detectable non-specific protease contaminating activities
Can be used under native or denaturing conditions
Glycerol-free for optimal performance in HPLC and mass spectrometry analysis
Supplied in solution; optimal activity and stability for up to 24 months
O-Glycoprotease is a highly specific protease that cleaves the peptide bonds of a glycoprotein or glycopeptide immediately N-terminal to a serine or threonine residue containing a mucin-type O-linked glycan with or without sialylation. O-Glycoprotease has a 6xHis-tag for easy removal from a reaction using nickel affinity resins.
Specificity:
Product Source
Cloned from Pseudomonas aeruginosa and expressed in E. coli.
Reagents Supplied
The following reagents are supplied with this product:
One unit of O-Glycoprotease will cleave > 90% of 2 µM FAM-labelled O-glycopeptide in a total reaction volume of 20 µl in 2 hours at 37°C in 20mM Tris-HCl, pH 8.0.
Reaction Conditions:
20mM Tris-HCl, pH 8.0.
Incubate at 37°C for 2 hours
Storage Buffer
20 mM Tris-HCl
100 mM NaCl
pH 7.5 @ 25°C
Heat Inactivation
95°C for 10 min
Molecular Weight
Theoretical: 97 kDa
Unit Assay Conditions
Two-fold dilutions of O-Glycoprotease are incubated with 2μM of FAM-labelled O-glycopeptide and 20 mM Tris-HCl pH 8.0 in a 20 µl reaction. The reaction mix is incubated at 37°C for 2 hours. Separation of reaction products are determined by CE analysis.
O-Glycoprotease cleaves a variety of glycoproteins and glycopeptides containing mucin-type O-glycans with or without sialic acid, including Tn-antigen (GalNAc-α-Ser/Thr).
Glycoproteins can be sequentially digested with O-Glycoprotease followed by Trypsin-ultra Mass Spectrometry Grade (NEB #P8101).
The optimal reaction buffer for O-Glycoprotease activity is 20 mM Tris-HCl, pH 8.0. The optimal pH range for O-Glycoprotease is pH 7.0-8.0. Enzyme activity decreases significantly at pH values below 7.0 and above pH 8.0.
Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.
The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]
The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]
This product is covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB).
While NEB develops and validates its products for various applications, the use of this product may require the buyer to obtain additional third party intellectual property rights for certain applications.
For more information about commercial rights, please contact NEB's Global Business Development team at gbd@neb.com.
This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.
New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.
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